Automatic air volume control system for ventilation of two patients using a single ventilator: a large animal model study

The COVID-19 pandemic outbreak led to a global ventilator shortage. Hence, various strategies for using a single ventilator to support multiple patients have been considered. A device called Ventil previously validated for independent lung ventilation was used in this study to evaluate its usability for shared ventilation. We performed experiments with a total number of 16 animals. Eight pairs of pigs were ventilated by a ventilator or anesthetic machine and by Ventil for up to 27 h. In one experiment, 200 ml of saline was introduced to one subject’s lungs to reduce their compliance. The experiments were analyzed in terms of arterial blood gases and respiratory parameters. In addition to the animal study, we performed a series of laboratory experiments with artificial lungs (ALs). The resistance and compliance of one AL (affected) were altered, while the tidal volume (TV) and peak pressure (Ppeak) in the second (unaffected) AL were analyzed. In addition, to assess the risk of transmission of pathogens between AL respiratory tracts, laboratory tests were performed using phantoms of virus particles. The physiological level of analyzed parameters in ventilated animals was maintained, except for CO2 tension, for which a permissive hypercapnia was indicated. Experiments did not lead to injuries in the animal’s lungs except for one subject, as indicated by CT scan analysis. In laboratory experiments, changes in TV and Ppeak in the unaffected AL were less than 11%, except for 2 cases where the TV change was 20%. No cross-contamination was found in simulations of pathogen transmission. We conclude that ventilation using Ventil can be considered safe in patients undergoing deep sedation without spontaneous breathing efforts.


Laboratory simulations of the Ventil device malfunction:
We performed tests to simulate a Ventil device malfunction when the function of automatic stabilization and division of the minute ventilation is abandoned. It covers the following situations: the electronic power failure, including a medical power supply unit failure, the temporary power grid failure in a medical facility, as well as the Ventil control electronic system failure. In a stable situation, we unplugged the power cord from the Ventil socket what immediately stopped the Ventil device. We made this "unplug" experiment in different respiratory stages, i.e., during the inspiration, end-inspiratory pause, and expiration. We repeated these experiments, but for the situation that powered off the Ventil device is powered on by plugging the power cord to the socket, simulating, for example, power grid restoration in the hospital. The results are presented in figures S9-S14. Additionally, we have successfully tested the Ventil device, for its continuous working, for two weeks.
The above results show that in a steady state when the Ventil stabilized division of the minute ventilation, the potential malfunction of the system could be safe for patients. The Ventil still divides the flow, and turning the power off and then on has virtually no effect on the flow and pressure signals. The Ventil dividing head is in a fixed position when the device is not powered; thus, it can still divide the flow but passively and potentially less accurate. However, in the case of the patient's respiratory improvement or deterioration, the position of the dividing head will not be changed until the power supply is restored. Therefore, in this case, the Ventil operates as a passive splitter.
Static pressure-flow characteristics 3 We determined the static inspiratory peak pressure drop (dPpeak)maximal flow characteristics for the Ventil device. For 1:1 flow division, the dPeak between the Ventil input and two outputs (dPeak1, dPeak2) were collected. The experiments were done for various minute ventilation values, respiratory rate values, and inspiratory to expiratory ratios as well. The characteristics are presented in figure S15. Flow Max in this figure is the maximal flow set in ventilator (volumecontrolled mode with rectangular inspiratory flow shape), while the dPeak1 is the difference between the inspiratory peak pressure obtained on the ventilator and measured in the Ventil left output channel and the dPeak2 is the difference between the same peak pressure from the ventilator and measured in the Ventil right output channel. We have obtained very linear characteristics within the measured range with a coefficient of determination R2 close to 1. We must point out that for higher flow rates, the differences between the pressure in the individual breathing circuit and the pressure measured by the ventilator can be up to 20 cmH2O for 1:1 ventilation. The peak pressures in the ventilator can be higher than typical and trigger too highpressure alarms. Pressure alarms specific management is required then due to elevated peak pressures in the ventilator.  Step changes (events) of the resistance (R) in the #1 object (artificial lung) on objects ventilation. Blue traces denote #1 ventilated object, while red traces denote #2 ventilated object.
"RX to RY" refers to the initial resistance (X) and the resistance after the event (Y), when the compliances are default (C=75) and constant; R values are expressed in mbar/L/s; Ventil device ratio is about 1:1; Black arrowa moment of the event. Respiratory rate RR = 12 1/min

Fig. S4.
Step changes (events) of the resistance (C) in the #1 object (artificial lung) on objects ventilation. Blue traces denote #1 ventilated object, while red traces denote #2 ventilated object.
"CA to CB" refers to the initial compliance (A) and the compliance after the event (B), when the resistances are default (R=5) and constant; "AL to Bag" refers to the exchanging the artificial lung (R=5, C=75) to the 2L respiratory bag (R~0, C~15); R values are expressed in mbar/L/s; C are expressed in ml/mbar; Ventil device ratio is about 1:1; Black arrowa moment of the event.

Fig. S5.
Step changes (events) of the resistance (R) in the #1 object (artificial lung) on objects ventilation. Blue traces denote #1 ventilated object, while red traces denote #2 ventilated object.
"RX to RY" refers to the initial resistance (X) and the resistance after the event (Y), when the compliances are default (C=75) and constant; R values are expressed in mbar/L/s; Ventil device ratio about 1:1; Black arrowa moment of the event. Respiratory rate RR = 18 1/min

Fig. S6.
Step changes (events) of the resistance (C) in the #1 object (artificial lung) on objects ventilation. Blue traces denote #1 ventilated object, while red traces denote #2 ventilated object.
"CA to CB" refers to the initial compliance (A) and the compliance after the event (B), when the resistances are default (R=5) and constant; "AL to Bag" refers to the exchanging the artificial lung (R=5, C=75) to the 2L respiratory bag (R~0, C~15); R values are expressed in mbar/L/s; C are expressed in ml/mbar; Ventil device ratio is about 1:1; Black arrowa moment of the event.

Fig. S7.
Step changes (events) of the resistance (R) in the #1 object (artificial lung) on objects ventilation. Blue traces denote #1 ventilated object, while red traces denote #2 ventilated object.
"RX to RY" refers to the initial resistance (X) and the resistance after the event (Y), when the compliances are default (C=75) and constant; R values are expressed in mbar/L/s; Ventil device ratio is about 1:1; Black arrowa moment of the event. Respiratory rate RR = 24 1/min

Fig. S8.
Step changes (events) of the resistance (C) in the #1 object (artificial lung) on objects ventilation. Blue traces denote #1 ventilated object, while red traces denote #2 ventilated object.
"CA to CB" refers to the initial compliance (A) and the compliance after the event (B), when the resistances are default (R=5) and constant; "AL to Bag" refers to the exchanging the artificial lung (R=5, C=75) to the 2L respiratory bag (R~0, C~15); R values are expressed in mbar/L/s; C are expressed in ml/mbar; Ventil device ratio is about 1:1; Black arrowa moment of the event.
Respiratory rate RR = 24 1/min     Technical report (IBBE PAS, 22-07-2020) -transmission of solutions/nanospheres in two respiratory branches of the Ventil system The aim of the research was to test the possibility of nanoparticle transmission between the two respiratory branches of the Ventil system. The research was carried out in three stages, respectively, with the use of (1) fluorescent nanopowders, (2) solutions of fluorescent compounds, and (3) suspension of the SARS-CoV-2 coronavirus phantom in the form of fluorescent nanospheres.
For some viruses, such as HIV, the term viral load is a numerical expression of the quantity of virus in a given volume of fluid in a sample volume, e.g. blood, plasma, etc. In contrast, for viruses that are transmitted by airborne droplets, a better term is called viral shedding. Moving on to studies related to infections caused by the droplet method, which are not many, one article quantified the presence of influenza A and B viruses in the exhaled air. The rate of increase in the number of influenza virus RNA particles on the mask filter has been found to be 3.2 to 20 particles (virions) per minute (can be directly related to the number of viral particles) [Fabian et al., 2008]. However, the data described in the above articles do not allow for the determination of the minimum infectious dose of virus that can be transmitted by inhalation. In the absence of data on SARS-CoV-2 infection, a model based on the influenza virus infection model was adopted. The influenza virus and SARS-CoV-2 are singlestranded RNA viruses of similar size -approx. 120 nm.
In studies using the viral phantom in the form of fluorescently labeled nanospheres, the estimated phantom titer was determined from an exponential pattern of viral reproduction in the respiratory system.
Moreover, according to the data presented in the publication [Chu et al., 2020], it was indicated that in the culture medium of Caco2 and Calu3 cells infected with SARS-CoV-2 virus, 120 h after infection, the number of virus genome copies was determined to be 9.5 . 10 9 in 1 mL of supernatant. The multiplicity of infection (MOI), defined as the ratio of the number of virus particles to the number of target cells present in these cultures, was 0.1.
Therefore, in our research the concentration of fluorescent nanospheres (viral phantoms) with a diameter of 100 nm was assumed at the level of approx. 24 . 10 12 particles per 3 mL, corresponding to the viral titer after incubation of the infected cell culture, e.g. Calu3 -human lung cancer cell line, cells over 5 days [Chu et al., 2020] with a 1000-fold allowance ensuring the possibility of optical evaluation of nanoparticle transmission in the ventilator tract.
Technical studies of the transmission of nanoparticles/solutions in two respiratory branches of the Ventil system were carried out in the period April 4 -29, 2020.

Chemicals and materials
Fluorescent compounds: sodium fluorescein (fluorescein) and methylene blue  Virus phantom: fluorescent red and green polystyrene nanospheres (NS) with a diameter of 100 nm. The stable fluorescence of the nanospheres was ensured by the incorporation of a dye inside them (so-called internal marking).

Testing solutions and suspensions
Fluorescein and methylene blue solutions with a concentration of 1 mM were used in the research. In contrast, fluorescent nanospheres were administered as a 60 µL suspension and 240 µL stock suspension (provided by the manufacturer) per 3 mL of deionized water. The estimated number of nanospheres in the above-mentioned the suspensions were 6.5 . 10 12 and 24 . 10 12 particles/3 mL. Figure R1 shows a diagram of the test set-up with the Ventil apparatus supplying the respiratory tract during independent ventilation of two artificial lungs (R and L). The measuring stand was prepared by the Ventil project team. Information on the settings of the ventilator and the associated Ventil is provided in Appendix 1. The L: R flow ratio was 6:4. In order to test the possibility of transmission between the airways, the test solutions/suspensions were administered to the expiratory tract of the artificial L lung using the Areogen nebulizer (N) in 3 mL portions repeated several times. The nebulizer operated in the mode of continuous and intensive nebulization -consumption of 5 mL solution within 30 minutes. The presence of test substances and nanoparticles was examined at test points T1, T2, and T3. Fluorescence observations of the respiratory tract elements, including filters, were carried out and documented with photos taken with a digital camera. The highlights of the respiratory tract were made with the use of a diode emitting UV radiation with a wavelength in the range of 360 nm.

Experimental measurement system/setup
Due to the fact that in the UV light autofluorescence of materials from which the elements of the measurement path are made is easily visible as a pink glow, attention should be paid to the selection of the colors of fluorescent substances used in the tests. After the pre-testing of visibility of the fluorescent dyes and nanospheres in the system, fluorescein and red fluorescent nanospheres were selected for the further study. In the case of using the electrostatic filter F in the position shown in Fig R1 and Fig. R2, typically used during artificial ventilation of the patient, no traces of fluorescent substances introduced into the system using a nebulizer were observed at the outlet of the F filter.
In the further part of the study, the F filter was applied from the artificial lung side upstream of the nebulizer or was not included in the system. Removal of filters from the system created favorable conditions for the transfer of test solutions and suspensions through the system. System without F electrostatic filter in the respiratory path with connected nebulizer containing fluorescein solution (Fig. R3). Fluorescein solution (yellowish green glow) is visible in the expiratory tract (right pipe). However, due to the overpressure in the inspiratory tract (left pipe), the fluorescein solution is not introduced there despite the lack of an F filter.
The respiratory tract filters were placed individually at the test points T1, T2, and T3. The surface of the filters at test points after exposure to four nebulization cycles of fluorescein solutions is shown in Figure R4. There were no traces of fluorescein on the post-filtering surfaces of the filters located in the test points. Furthermore, no traces of fluorescein were observed on the filtering surface of the filter placed at test point T3. After exposure to 4 nebulization cycles, traces of the suspension of fluorescent nanospheres were observed only on the filter surface of the filter placed at test point T1 (Fig. R6. A and B). At the T2 test point, traces of nanospheres were found on the filtration surface (no photo was included due to poor quality). However, at the last test point T3, no nanoparticles were found on the filtration surface.

Summary
The report presents the results of technical studies on the transmission of nanoparticles / solutions in two respiratory branches of the Ventil system. The experiment conditions were selected in such a way that they were unfavorable from the point of view of protection against transmissionintroducing individual filters into the system at subsequent test points. Traces of fluorescein on the filtering surface of the filter at test point T3 (ventilator entry) only occur with fluorescein solution. On the other hand, in the case of fluorescent nanoparticles introduced from the suspension with an excess concentration (approx. 3 orders of magnitude), they appear on the filtering surface of the filter at point T1. On the basis of the conducted technical studies related to the assessment of the possibility of transmission of solutions/suspensions in two respiratory branches of the Ventil system, it was found that under the conditions of the experiment with the use of fluorescent solutions and suspensions no transmission occurs. To complement, based on the research it was found that there was no transmission of phantoms of virus -fluorescent nanospheres of 100 nm diameter, between the two pathways of the respiratory system during independent ventilation of two artificial lungs using the Ventil apparatus.
The independent studies described in this report, in some part related to the use of solutions of fluorescent compounds to evaluate the possibility of transmission, were conducted under similar conditions and are consistent with the results published in the article [Srinivasan et al. 2020]. In relation to the research described in the above-mentioned paper (continuous nebulization of a 5 mL solution of trypan blue for 10 minutes, and visual observation of filters), in our experiment, apart from use fluorescein and methylene blue solutions, we proposed a method of examination of transmission, which used phantoms of virus in the form of internally labeled with fluorescent dyes nanospheres with a diameter of 100 nm, the concentration of which in the administered suspension was approx. 24 . 10 12 particles/3 mL, corresponding to the viral titer with ca. 1000-fold excess, in respect to SARS-CoV-2 titer after incubation of the infected Calu3 cell culture for more than 5 days [Chu et al., 2020]. Fluorescent solutions and suspensions were introduced into the respiratory tract in 4 continuous nebulization cycles lasting 30 minutes each. Research with the use of nanoparticles can be carried out in laboratory environment, giving preliminary information about the possibility of cross-transmission in the respiratory tract during independent ventilation of artificial lungs.